prevalence, genetic diversity, and antimicrobial resistance patterns of arcobacter and ...

133 Pages·2005·0.78 MB·English

PREVALENCE, GENETIC DIVERSITY, AND ANTIMICROBIAL RESISTANCE
PATTERNS OF ARCOBACTER AND CAMPYLOBACTER ON BROILER CARCASSES
DURING PROCESSING
by
INSOOK SON
(Under the Direction of Mark A. Harrison)
ABSTRACT
Broiler carcasses (n = 325) were sampled in a U.S. commercial poultry processing plant during
five plant visits from August to October of 2004 at three sites along the processing line: 1) pre-scalding,
2) pre-chilling, and 3) post-chilling. Arcobacter species were recovered from pre-scalded carcasses more
frequently (96.8%) than from pre-chilled (61.3%) and post-chilled carcasses (9.6%). For Arcobacter
identification, a species-specific multiplex PCR assay showed that A. butzleri was the most prevalent
species (79.1%) followed by A. cryaerophilus 1B (18.6%). A. cryaerophilus 1A was found at low levels
(2.3%) and A. skirrowii was not isolated at all. Campylobacter was isolated from 92% of pre-scalded
carcasses, 100% of pre-chilled carcasses, and 52% of post-chilled carcasses. For Campylobacter
speciation, the BAX® PCR identified as C. jejuni (87.6%) as the most common species followed by C.
coli (12.4%).
The genetic diversity of Arcobacter and Campylobacter was analyzed by pulsed field gel
electrophoresis (PFGE). Genomic DNA was digested with KpnI from Arcobacter strains and SmaI from
Campylobacter strains. A total of 32.8% of Arcobacter isolates belonged to single-isolate groups, while
only 2.3% of Campylobacter isolates belonged to this category. The remaining Arcobacter species were
distributed among 25 multi-isolate PFGE groups, while Campylobacter species were found in just eight
multi-isolate groups.
The great majority of Arcobacter (93.7%) and Campylobacter (99.5%) isolates were resistant to
one or more antimicrobials. Multiple antimicrobial resistance was observed in 71.8% of the Arcobacter
isolates and in 28.4% of the Campylobacter isolates. Of the A. butzleri isolates, 89.9% (n = 125) were
resistant to clindamycin, 82% (n = 114) were resistance to azithromycin, and 23.7% (n = 33) were
resistant to nalidixic acid. Resistance to tetracycline was very high in C. jejuni and C. coli at 99.5% and
96.3%, respectively.
These data suggest significant contamination of Arcobacter and Campylobacter from carcasses
from different processing sites in a commercial poultry plant with a high genetic diversity of Arcobacter,
and demonstrated resistance in Arcobacter and Campylobacter to common antimicrobial agents.

INDEX WORDS: Arcobacter, Broiler chickens, Campylobacter, Poultry processing, Genetic
diversity, PFGE, Typing, Antimicrobial resistance
PREVALENCE, GENETIC DIVERSITY, AND ANTIMICROBIAL RESISTANCE
PATTERNS OF ARCOBACTER AND CAMPYLOBACTER ON BROILER CARCASSES
DURING PROCESSING

by

INSOOK SON
B.S., Kyungpook National University, Korea, 1997
M.S., Kyungpook National University, Korea, 1999

A Dissertation Submitted to the Graduate Faculty of The University of Georgia in Partial
Fulfillment of the Requirements for the Degree

DOCTOR OF PHILOSOPHY

ATHENS, GEORGIA
2005
© 2005
Insook Son
All Rights Reserved
PREVALENCE, GENETIC DIVERSITY, AND ANTIMICROBIAL RESISTANCE
PATTERNS OF ARCOBACTER AND CAMPYLOBACTER ON BROILER CARCASSES
DURING PROCESSING

by

INSOOK SON

Major Professor: Mark A. Harrison

Committee: Paula J. Fedorka-Cray
Mark D. Englen
Joseph F. Frank
Larry R. Beuchat

Electronic Version Approved:

Maureen Grasso
Dean of the Graduate School
The University of Georgia
December 2005
DEDICATION

This dissertation is dedicated to
My parents
And
My parents-in-law
For their prayer, constant support, and unconditional love.

AND
To
My beloved husband
Kwangwook Ahn
For his unwavering encouragement, support, and patience.

Without all of you, I could never have come so far.
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ACKNOWLEDGEMENTS

I would like to acknowledge my major professor, Dr. Mark A. Harrison, and co-advisors, Dr.
Mark D. Englen and Dr. Paula J. Fedorka-Cray for their invaluable guidance, constant encouragement,
and financial support throughout my Ph. D. program. I would also like to acknowledge Dr. Mark E.
Berrang for his wealth of knowledge and willingness to help at any time. I am very grateful to Dr. Joseph
F. Frank and Dr. Larry R. Beuchat for serving on my advisory committee. I would like to express special
thanks to Scientist Scott R. Ladley for his invaluable advice and encouragement. I like also to thank to
Mark N. Freeman for assistance with sampling and kindness. Appreciation is extended to the many
members of the Bacterial Epidemiology and Antimicrobial Resistance Research Unit at the Russell
Research Center and friends in the Department of Food Science and Technology for all their friendship
and invaluable help.
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TABLE OF CONTENTS
Page
ACKNOWLEDGEMENTS.............................................................................................................v
LIST OF TABLES........................................................................................................................vii
LIST OF FIGURES.....................................................................................................................viii
CHAPTER
1 INTRODUCTION.........................................................................................................1
2 LITERATURE REVIEW..............................................................................................7
Taxonomic history and general characteristics.........................................................7
Sources......................................................................................................................9
Arcobacter, Campylobacter and human infection..................................................12
Pathogenesis............................................................................................................13
Isolation and identification methods.......................................................................15
Typing methods for epidemiological studies..........................................................19
Antimicrobial susceptibility of Arcobacter and Campylobacter............................23
3 PREVALENCE OF ARCOBACTER AND CAMPYLOBACTER ON BROILER
CARCASSES DURING PROCESSING................................................................58
4 GENETIC DIVERSITY OF ARCOBACTER AND CAMPYLOBACTER ON
BROILER CARCASSES DURING PROCESSING..............................................79
5 ANTIMICROBIAL RESISTANCE PATTERNS OF ARCOBACTER AND
CAMPYLOBACTER ON BROILER CARCASSES DURING PROCESSING...100
6 SUMMARY AND CONCLUSIONS........................................................................121
vi
LIST OF TABLES
Page
Table 3.1: Arcobacter on broiler carcasses from poultry processing plant...................................73
Table 3.2: Arcobacter species on broiler carcasses from poultry processing plant.......................74
Table 3.3: Campylobacter on broiler carcasses from poultry processing plant.............................75
Table 3.4: Campylobacter species on broiler carcasses from poultry processing plant................76
Table 4.1: Distribution of PFGE patterns of Arcobacter on broiler carcasses from poultry
processing plant............................................................................................................95
Table 4.2: Distribution of PFGE patterns of Campylobacter on broiler carcasses from poultry
processing plant...........................................................................................................96
Table 5.1: Percentage of Arcobacter isolates on broiler carcasses resistant to antimicrobials by
species........................................................................................................................117
Table 5.2: Percentage of Campylobacter isolates on broiler carcasses resistant to antimicrobials
by species...................................................................................................................118
Table 5.3: Resistance patterns of Arcobacter isolates on broiler carcasses resistant to two or more
antimicrobials.............................................................................................................119
Table 5.4: Resistance patterns of Campylobacter isolates on broiler carcasses resistant to two or
more antimicrobials...................................................................................................120

vii
LIST OF FIGURES
Page
Figure 3.1: Species-specific multiplex PCR of Arcobacter species..............................................77
Figure 3.2: Species-specific BAX® PCR of Campylobacter species............................................78
Figure 4.1: (A) PFGE KpnІ restriction profiles of Arcobacter isolates: Lanes 1-6 and 10-11, pre-
scalding; Lanes 7-9, post-chilling. Lane M, Salmonella Braenderup H9812
molecular size standard. (B) PFGE SmaІ restriction profiles of Campylobacter
isolates: Lanes 1-7 and 9-10, pre-scalding; Lane 8, post-chilling. Lane M, Salmonella
Braenderup H9812 molecular size standard................................................................97
Figure 4.2: PFGE patterns of the A. butzleri (B-1 to B-20), A. cryaerophilus 1A (C.1a), and A.
cryaerophilus 1B (C.1b-1 to C.1b-3) isolates. Collections site: 1, pre-scalding; 2, pre-
chilling; 3, post-chilling. The dendogram was generated using UPGMA cluster
analysis and Dice similarity coefficient......................................................................98
Figure 4.3: PFGE patterns of the C. coli (C-1) and C. jejuni (J-1 to J-7) isolates. Collections site:
1, pre-scalding; 2, pre-chilling; 3, post-chilling. The dendogram was generated using
UPGMA cluster analysis and Dice similarity coefficient........................................... 99

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